The invention relates to a measurement assembly for parallel readout of surface plasmon resonance (SPR) sensors.
In the search for new active substances combinatorial chemical systems hold high promise in finding ligands matching a receptor molecule. Miniaturizing and automating synthesis and parallelization thereof is salient to assaying as large a number of ligands as possible. Due to the small resulting amounts of ligand, these requirements (miniaturizing, automating and parallelizing) apply likewise to detecting the ligand receptor binding for which the high sensitivity of the SPR method can be used in which the light reflected from a thin gold film is detected. Under a suitable resonance condition (angle of incidence and wavelength of the light and thickness of the gold film) the intensity of the reflected light is reduced. The energy of the light is then transformed into charge density waves of the electron gas in the gold film, these charge density waves being termed plasmons.
There are two methodical approaches to observing the resonance: either monochromatic light is used in plotting the intensity of the reflected light as a function of the angle of incidence or the angle of incidence is maintained constant and the wavelength of the light is varied. In both cases the resonance curve is shifted with a change in the refractive index of the medium provided on the side of the gold film facing away from incident light.
This effect is made use of in biochemical analysis. The receptor or ligand is immobilized on the gold surface. After addition of the ligand or receptor the resonance condition is changed on molecular association.
The simplest assembly for measuring this effect is a glass prism which is illuminated with light and the angle of incidence of which is varied (see e.g. xe2x80x9cBiospecific interaction analysis using biosensor technologyxe2x80x9d Malmqvist, M., Nature 361 (1993) 186-187).
A more sophisticated method is the parallel detection of multiple angles in which the gold surface is illuminated with a slightly divergent beam of monochromatic light (aperture angle xcx9c10xc2x0) and the reflected light is directed to a position resolving light detector for obtaining an explicit assignment between the angle of reflection and the position on the detector. This construction has the advantage of sensing the angle range of interest with no moving parts. This is why use is made of this kind of detection in a few commercial systems as disclosed e.g. in WO 90/05295 A1 or EP 797 091 A1. One disadvantage of these assemblies is that only one prepared array of gold sensors (one-dimensional detection) or but a few arrays of sensors arranged along a line (using a two-dimensional detection) can be assayed each time, i.e. this not permitting simultaneous measurement of a two-dimensional sensor array by this angle detection method. After having installed the prepared gold film in systems of this kind, obtaining thermal equilibrium takes, however, some minutes (at least 15 minutes), i.e. the actual measurement then lasting at least until equilibrium of the molecular association is attained which may also take up some minutes. This is why systems of this kind lack good suitability in detecting the binding of a large number of different ligands, since the time and expense involved in measuring and changing the samples is relatively high.
A parallel approach to analyzing a sample array is SPR microscopy (SPM) (see: EP 388 874 A2 or M. Zizisperger, W. Knoll, Prog Colloid Polym Sci. 1998, 109 pages 244-253) in which the gold surface applied to a prism is coated in various portions with various samples and the gold surface imaged on a CCD chip at the SPR angle. During measurement the angle is varied by a mechanical scanner. However, this method is restricted to small object diameters.
A more recent SPR method is disclosed in WO 94/16312A1 in which detecting the binding of small amounts of substance is achieved by optical fiber guides partially coated with a gold film. However, here too, the problem still exists in designing a system required to assay a plurality of samples in parallel in accordance with this principle. Such an array of gold-coated fibers is, on the one hand, expensive and highly sensitive to mechanical stress, and, on the other, producing the array in parallel as proposed therein is difficult to achieve technically.
Optical fiber guides are also employed as it reads from WO 98/32002 A1. To protect them from being damaged physically the fiber cable is housed in a pipette. To achieve an array it is proposed to use a series arrangement of such pipettes. However, miniaturizing such an arrangement is difficult to achieve, especially for parallel measurement of many different samples.
Known from DE 196 15 366 A1 is a method and a means for the simultaneous detection of physical, biological or biochemical reactions and interactions at or in the surface of samples. All samples are simultaneously illuminated with monochromatic light and the reflected light supplied to a detection array, e.g. of a CCD matrix or video camera. The samples may be matrixed with the sensor surface areas in a single plane on a substrate plate located in turn on a transparent base plate which is illuminated. The electrical signals of the detection array are supplied to an analyzer. After analyzing the samples at one wavelength, the wavelength can be set to another value.
Known from WO 95/22754 is an analyzer in which a plurality of SPR analysis wells are arranged in a matrix for simultaneous assaying. The sensor surface areas of the analysis wells are located parallel in a single plane.
The invention is based on the object of defining a measurement assembly for simultaneous readout of a plurality of SPR sensors, more particularly exceeding a hundred or a thousand, in which readout is completed with a measuring time of less than thirty minutes.
This object is achieved by the measurement assembly and the measurement method as set forth in the independent claims, advantage aspects reading from the sub-claims.
To achieve this object, use is made of a specially configured array in an assembly with imaging methods to permit simultaneous readout. The array used for this purpose comprises a plurality of waveguides, it being noted that waveguide in this context is understood within the scope of the present invention to be an optical medium in which the light is guided in at least one dimension and which has at least two parallel interfaces.